TW201505670A - Matrix for the infiltration with cells - Google Patents

Abstract

A cell implant matrix has a connective porosity of more than 80 % and consists mainly of a mixture of bioresorbable polymers, wherein the matrix has disk-shape and wherein a surface layer on one side of the disk has less than 20 % of the average pore density of the other sides. The matrix is manufactured by providing a bioresorbable polymer layer; stratifying, onto the polymer layer, a mixture of a water-soluble solid, at least two polymers differing with respect to their resorption rates, and a solvent for one of the polymers; evaporating the solvent optionally followed by compacting the mixture; and watering the compacted body to remove the salt.

Description

Cell infiltration matrix

This application is a porous substrate for cell penetration for therapeutic or diagnostic use.

Cell implantation based on a porous matrix made of a biocompatible polymer is known from WO 2004/108810 A1. In such matrices, the pores are connected and have the function of a template for in vivo cell permeation (e.g., treatment) or in vitro cell permeation (e.g., diagnosis). For transplantation, such bioresorbable matrices have the function of temporarily localizing the implant.

In some applications, known templates are not yet fully satisfactory, especially for their clinical efficacy.

It is an object of the invention to improve the clinical efficacy of a template.

To this end, the present invention provides a conventional dish-shaped template in which the average pore amount of one surface on one side of the template is less than 20% of the average pore amount on the other side (complex side). This asymmetric structure allows the template to be placed into the body in such a way that the implantable viable cells remain in the template for a longer period of time.

According to another aspect, the present invention provides a method of producing a porous bioresorbable substrate, wherein: initially forming a polymer layer having no pore former; forming at least two polymers on the polymer layer a solvent for one of the polymers, and a mixture of water-soluble pore formers; the solvent is then evaporated and then watered to remove the pore former. In a variant, pressurization can be carried out between the next two steps. Both methods can obtain a highly porous polymeric matrix disc, but have no pores on one side or at least a film with less pores. Despite this film, the cell supply present in the pores during use is still sufficient, but it is greatly reduced because The rate of loss caused by the migration. In a variation of the template having the function of supporting a connective tissue, the continuous polymer layer can provide sufficient strength for, for example, a suture material for a fixed template.

In the examples, the porous substrate is hydrophilically coated with, for example, polymerizable (meth)acrylic acid. To this end, a plasma coating step is followed by a plasmaless coating step whereby a desired film thickness greater than one micron is achieved.

Further features of the present invention are provided in the following description of the embodiments and the claims and claims. The invention is not defined by the described embodiments, but is defined by the scope of the appended claims. In particular, independent features of embodiments of the invention may be implemented in numbers and combinations other than the following examples. In the explanation of the following embodiments, reference is made to the accompanying drawings showing a flow chart of the method according to the invention.

S1‧‧‧Manufacture of polymer layers with no or less pores

S3‧‧‧ A mixture of two polymers, a solid pore former, and a solvent for the polymer

S5‧‧‧ Evaporation solvent

S7‧‧‧ watering to remove pore formers

Figure 1 is a flow chart of a method in accordance with the present invention.

In a major application, a matrix for implanting functional cells, such as hepatocytes and/or islet cells, is provided. Such biochemically functioning cells adhere to the inner walls of the pores of the foamy matrix (adhesion rate exceeds 80%, or over 95% when properly coated) and ideally can be grafted to the cells along with the matrix The donor's own cell mesothelial pockets. It has been found here that no rejection occurs in this case but only a slight foreign matter is stimulated, which is especially advantageous for therapeutic treatment. Within a few weeks, the matrix is vascularized and the implanted cells no longer rely solely on diffusive supplies. The matrix is specifically configured such that less pores (or no pores) are laterally inward and more pores are outwardly outward to maintain a low level of loss due to migration. At the same time, a portion of the resorbable matrix gradually reabsorbs (within 3-4 months, or at least 2 and/or less than 7 months) and the physiological environment is thereby affected to similarly favor treatment success. It is generally desirable that a portion of the polymer mixture be more slowly corroded (the ratio of degradation time is at least 5) and maintain structural integrity for extended periods of time, such as 2.5-3 years (or at least 2 years and/or less than 5 years). Such polymers are preferably based on alpha-hydroxycarbonic acids such as lactic acid and/or glycolic acids (such as PLA or PLGA). Manufacturers of such polymers that are certified for use in the human body point to nominal degradation times that are of great significance here.

As noted above, a particularly good adhesion rate was observed on the coated substrate, initially in a combined treatment of PECVD/CVD by a thin PAA-layer (eg 20-30 nm) plasma coating followed by no plasma. A substrate coated with a thicker PAA-layer (eg, 20-30 μm) under action. This upper layer forms a crystalline hydrophilic layer.

Initially, a solution of one of the polymers used for medical use certification in chloroform was poured into a mold, and the solvent was evaporated at a temperature of 45 to 65 °C. Next, a polymer mixture having a predetermined particle size distribution is mixed with a rock salt particle similarly having a predetermined particle size distribution, and then mixed with a solution of one of the polymers in chloroform, and then the above mixture is brought to an existing one. Above the polymer layer. The solvent is evaporated from the preform at a slightly elevated temperature (45 ° C - 65 ° C) and pressurized if necessary to make it compact. This preform is watered to remove salts, thereby providing the desired porosity. However, in this paper, the polymer layer initially manufactured remains unpored. Depending on the field of use, the thickness of the film layer with less porosity can be controlled by the amount and concentration of the initial solution. For example, if the concentration of the solution is low (for example, 4% of the slowly degradable polymer in chloroform) and the degree of filling is small (for example, 0.1-1 mm such as 0.3 mm), an extremely thin film can be obtained. When it is desired to obtain a structure with a high mechanical tolerance, the degree of filling can be set higher at the same concentration (for example, 5-50 mm, usually 20-25 mm). In this case, evaporation of chloroform takes a long time (1.5 hours). In the former case, the obtained film has a thickness of about 10 to 20 μm, and in the latter case, the resulting film has a thickness of about 0.5 to 1 mm.

The layered mixture of rock salt particles (median falling at 350-370 μm) is lower than the polymer particles (the median of the more slowly degradable polymer falls between 210 μm and 230 μm, and the median of the polymer that can be more rapidly degraded falls at 150 μm Slightly thicker to between 170 μm. The distribution width (5%/95%) of the salt or total polymer is similar, i.e., about ±85-95 μm. The shape of the distribution may be bimodal or trimodal. The composition of the layered mixture is about 96% salt, 1-1.5% solid polymer, and about 3-5% dissolved polymer, wherein the volume ratio of solid to liquid is about equal. In total, the portion of the polymer that degrades faster is only about 5-20% of the polymer. The total thickness of the pore-forming layer is 5-6 mm. In variations with a more stable initial layer, a slightly coarser salt (median between about 400-420 μm) can be selected. In this case, the total thickness of the pore-forming layer is 4 to 5 mm and the step of pressurizing and solidifying can be dispensed with. The watering is continued for about 24 hours and then dried at a temperature of 45-50 °C. When the coating is formed, the matrix is disposed on the side with less voids, with the predominantly open open pore side.

The polymers used herein are, for example, those sold by Evonik, the names of which are These are L210s, L210, L09s, L207s, L206s (PLGA-polymers that can be slowly degraded) or RG502, RG502H, RG505 (PLGA-polymers that can degrade faster).

In in vitro applications, the matrix according to the invention may have the function of immobilizing cells of the agent that will be exposed to the bioreactor. For example, in this manner, it can be investigated whether the selected cell type will respond to the drug being studied, and the treatment can be planned based on the observations obtained thereby. Similarly, since any toxicity is recognized at an early stage, the development of the drug can be simplified.

S1‧‧‧Manufacture of polymer layers with no or less pores

S3‧‧‧ A mixture of two polymers, a solid pore former, and a solvent for the polymer

S5‧‧‧ Evaporation solvent

S7‧‧‧ watering to remove pore formers

Claims (19)

A substrate for cell implantation, the matrix having a porosity of more than 80% and consisting essentially of a mixture of bioabsorbable plurality of polymers, wherein the substrate is dished and characterized by: one side of the dish substrate The average pore ratio of the upper surface is less than 20% of the average pore ratio of the other side of the dish. A substrate for cell implantation according to the first aspect of the invention, wherein the substrate is coated with a hydrophilic coating. A substrate for cell implantation according to claim 2, wherein the coating is mainly composed of a (meth)acrylic unit. A substrate for cell implantation according to claim 2 or 3, wherein the thickness of the coating is greater than 1 μm. The substrate for cell implantation according to any one of claims 1 to 4, wherein the matrix is mainly composed of poly(α-hydroxy)carbonic acid. The substrate for cell implantation according to any one of claims 1 to 5, wherein the two polymers in the plurality of polymers constituting the mixture have a resorption rate different by more than 5 times, the two polymers Each of the polymers is at least 10% of the mixture. A method of manufacturing a substrate for cell implantation and having different interconnected porosity, the method comprising: fabricating a bioabsorbable polymer layer; unilaterally imparting a water-soluble solid with different resorption rates on the polymer layer Forming a film layer of at least two polymers, a mixture with at least one of the polymers, which is immiscible with water; evaporating the solvent; and watering the mixture after evaporation of the solvent to form a film therefrom The layer removes the water soluble solid. The method for producing a substrate for cell implantation and having different interconnected porosity, as in claim 7, further comprising applying a pressure after evaporating the solvent to make the mixture strong. A method of producing a substrate for cell implantation and having different connectivity porosity as in the seventh or eighth aspect of the invention, wherein chloroform is used as the solvent. The method for producing a substrate for cell implantation and having different connectivity porosity according to any one of claims 7 to 9, wherein a resorption rate of the two polymers in the polymer constituting the mixture differs by more than Five times, and each of the two polymers comprises at least 10% of the mixture. The method for producing a substrate for cell implantation and having different connectivity porosity according to any one of claims 7 to 10, further comprising coating with a hydrophilic component after the watering step. A method for producing a substrate for cell implantation and having different connectivity porosity according to the invention of claim 11, wherein the hydrophilic component is mainly composed of (meth)acrylic acid or (meth)acrylic anhydride. A method for producing a substrate for cell implantation and having different connectivity porosity according to claim 11 or 12, wherein a plasma coating step is carried out in the presence of a noble gas or an inert gas, and then plasma-free coating is carried out. Cloth steps. The method for producing a substrate for cell implantation and having different interconnected porosity according to any one of claims 7 to 13, wherein the polymer layer initially manufactured has a thickness of 0.002-2.5 mm. A method of producing a substrate for cell implantation and having different interconnected porosity according to claim 14 wherein the thickness of the polymer layer initially produced is 0.005 to 0.025 mm. A method of producing a substrate for cell implantation and having different interconnected porosity according to claim 14 of the invention, wherein the polymer layer initially produced has a thickness of 0.25 to 2.0 mm. The method for producing a substrate for cell implantation and having different connectivity porosity according to any one of claims 7 to 16, wherein the implant substrate produced is as claimed in claims 1 to 6. A substrate for cell implantation. The use of a substrate for cell implantation according to any one of claims 1 to 6 or a substrate produced by the method of manufacture according to any one of claims 7 to 17 for use in Infiltration of viable cells and use to expose the viable cells to a predetermined test agent in vitro. The use of a substrate for cell implantation according to any one of claims 1 to 6 or a substrate produced by the method of manufacture according to any one of claims 7 to 17 for treatment Infiltration of cells in the treatment of the human body.